chapter 15
Carbohydrate Metabolism II: Gluconeogenesis, Glycogen Synthesis and Breakdown, and Alternative Pathways
Other proteins involved in glycogen synthase control
are calmodulin, inhibitor-1, and inhibitor-2. Troponin C,
when complexed with Ca2+, can activate phosphorylase
kinase, but phosphorylase kinase may have little or no
role in the inactivation of glycogen synthase
in vivo.
Inhibitor-1 (M.W. 18,600), when phosphorylated on thre-
onine 35 by cAMP-dependent protein kinase, binds to and
inhibits protein phosphatase-1. The concentration of pro-
tein phosphatase
- 1
is lower than that of inhibitors, and its
activity is sensitive to the latter, suggesting that inhibitor
- 1
phosphorylation may be an important regulatory mech-
anism for this phosphatase. Inhibitor-2 (M.W. 30,500)
appears to be an integral part of protein phosphatase-
1, forming an inactive 1:1 complex, called Mg
dependent phosphatase, with the catalytic subunit of this
enzyme. Phosphorylation of inhibitor-2 (also at a threo-
nine residue) activates protein phosphatase-1. This phos-
phorylation, which requires Mg
+-ATP, is catalyzed by an
“activating factor” (F
a ),
Control o f Glycogen Phosphorylase
Regulation of glycogen phosphorylase in muscle is ac-
complished by many of the same enzymes that control
glycogen synthesis. Phosphorylase kinase converts the
dimeric phosphorylase from the inactive to the active form
by Mg2+ and ATP-dependent phosphorylation of two iden-
tical serine residues. The principal enzyme that removes
this phosphate may be protein phosphatase
- 1
lase phosphatase).
Phosphorylase kinase is a hexadecamer having four dif-
ferent subunits: a (M.W. 145,000) ora'(M.W. 133,000),
(M.W. 128,000),
(M.W. 44,700), and 5 (M.W. 16,700),
with the stoichiometry a 4(or
a'A)PAy 48^
(Figure 15-10).
Whether the
or a'-subunit is present depends on the
tissue. The subunits apparently differ in Ca2+ sensitivity,
since the 5'-subunit, discussed later, will not bind to the
A catalytic site on the y-subunit has considerable ho-
mology with the catalytic subunit of cAMP-dependent
protein kinase (Chapter 30). Evidence for active sites on
and /
-subunits is weak.
Phosphorylase kinase activity has an absolute require-
ment for Ca2+, which binds to the 5-subunit. The amino
acid sequence of this subunit is nearly identical to that of
calmodulin, with four calcium binding sites, but unlike
calmodulin, the 5-subunit is an integral part of the enzyme
and does not dissociate from it in the absence of Ca2+. In
the presence of Ca2+, kinase activity is further increased
by phosphorylation of the
and /
-subunits, catalyzed by
cAMP-dependent protein kinase and several other kinases.
Phosphorylation may activate the enzyme by disinhibiting
FIGURE 15-10
Subunit structure of muscle phosphorylase kinase. The
a -
and A-subunits
are regulatory proteins containing the sites at which phosphorylation can
activate the kinase. The //-subunit contains the catalytic site. The 5-subunit
is a Ca2+-binding protein, essentially identical with calmodulin, Ca2+.
Calcium binding to the 5-subunit causes a change in the quaternary
structure of the enzyme, allowing binding of the 5'-subunit, thereby
increasing kinase activity even in the absence of phosphorylation of the
A-subunit. [Modified and reproduced with permission from
A. K. Campbell:
In te rce llu la r C alcium : Its U n iversa l R o le a s Regulator,
Wiley, New York, 1983. © 1983 by John Wiley & Sons, Inc.]
it, since limited proteolysis (which degrades the
-subunits) and dissociation of the holoenzyme into par-
tial complexes
also cause activation. Acti-
vation by covalent modification increases the affinity of
the kinase for phosphorylase b. The kinase also is acti-
vated by the binding of a 5'-subunit (Ca2+/calmodulin or
Ca2+/troponin C) to a site formed by the
and /
There are sites for four 5'-subunits on the holoenzyme. Ac-
tivation by Ca2+/troponin C is 20-30 times greater than
by Ca
+/calmodulin. This route may provide a means of
coordinating muscle contraction, initiated by binding of
Ca2+ to troponin C (Chapter 21), with glycogenolysis.
Finally, phosphorylase kinase is allosterically activated by
the binding of up to
mol of ADP.
Only 20-30% of isolated phosphorylase kinase is as-
sociated with glycogen particles, whereas all glycogen
phosphorylase activity is associated with glycogen par-
ticles. This fact, together with the subcellular localiza-
tion of phosphorylase kinase in intact cells and its bio-
chemical properties, suggests that phosphorylase kinase
may control other metabolic pathways as well. Suggested
phosphorylase kinase targets
include the Na+,
and Ca2+-ATPases in the sacrolemma and sarcoplasmic
reticulum, respectively.
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